The effects of hydrogen peroxide, which fetal bovine serum (FBS) releases, on proliferation have been studied in ROS 17/2.8 osteoblasts. Cell proliferation, when activated by FBS, was inhibited by catalase in ROS 17/2.8 osteoblasts, but did not in primary osteoblast-like cells. Serum-induced extracellular signal-regulated kinase(ERK) activity was reduced by the pretreated catalase in ROS 17/2.8 osteoblasts. In addition, the present studies demonstrate that addition of FBS led to an increase of fluorescence of dihydrorhodamine 123, indicating formation of free radicals including hydrogen peroxide in ROS 17/2.8 osteoblasts, but not in primary osteoblast-like cells. These phenomena may account for the generation of reactive oxygen species during cellular proliferation in ROS 17/2.8 osteoblasts.